Despite being a staple therapy for pancreatic cancer (PC), Radiation Therapy (RT) provides limited objective clinical response due to inherently high radioresistance (RR) of PC. As the risk of radiation-induced toxicity for PC patients far outweighs the therapeutic benefits attained, effective methods to improve the radiosensitivity of PC are urgently needed. The overall objective of this project is to identify and characterize pathway(s) contributing to RR in PC that can be explored as novel targets for radiosensitization (RS). Preliminary global gene expression analysis suggested novel involvement of cholesterol biosynthesis pathway in the RR in PC cells. Inhibition of cholesterol biosynthesis (CBS) by Zoledronic acid (Zometa) resulted in RS of panel of RR murine and human PC cells. Further, this RS was recapitulated by the inhibition of the small GTPase Rac1, whose activity is controlled by the CBS pathway. Therefore, we seek to delineate mechanisms of RR mediated by cholesterol biosynthesis pathway, and evaluate the potential of Zometa as a radiosensitizer (RST) in preclinical and clinical studies. We propose to exploit the strength of stereotactic radiation therapy and genetically engineered mouse models to comprehensively test the hypothesis that cholesterol biosynthesis pathway contributes to radioresistance in PC and Zometa inhibits specific pathways consistently implicated in RR and its use will radiosensitize PC both in vitro and in vivo. To achieve our goal, three specific aims are proposed. Aim 1 will elucidate the mechanisms of radioresistance in PC and validation of FDPS/Rac1 inhibitors (Zometa and NSC23766) as RSTs. The functional role of critical genes identified will be determined by knockdown and overexpression studies, use of specific inhibitors, and immunohistochemistry on clinical samples. Aim 2 will determine the efficacy of Zometa as RST in mouse models (xenograft and autochthonous). A novel strategy of stereotactic irradiation for murine models will be developed. In Aim 3, a Phase-I/II study will be undertaken to assess the radiosensitizing potential of Zometa in human subjects and determine if Zometa is well tolerated in PC patients undergoing RT.